The genetic heterogeneity among Mycobacterium tuberculosis isolates from 501 patients in Ethiopia, Tunisia, and the Netherlands was compared by analysis of DNA polymorphism driven by insertion element IS6110. The percentage of isolates displaying two or more identical patterns differed greatly in the three countries: It was highest among Tunisian isolates and lowest in Dutch isolates. In contrast to isolates from Dutch subjects infected with M. tuberculosis, the majority of strains from Ethiopia and Tunisia were from a few families of genetically highly related strains. Furthermore, little overlap was observed among isolates from the three countries, indicating strict isolation of the bacterial reservoirs in the countries. A few strains from the Netherlands matched strains from Ethiopia and Tunisia. Those strains were invariably isolated from refugees, immigrants, or persons who visited Ethiopia or Tunisia.

In this study the polymerase chain reaction (PCR) with previously developed oligonucleotide primers was used to detect Leishmania aethiopica in paraffin-embedded skin biopsy specimens. The Leishmania-specific 120 base pair fragment of the kinetoplast deoxyribonucleic acid (kDNA) minicircles has been amplified from all parasitologically or histologically confirmed cases of cutaneous leishmaniasis (CL), as demonstrated by gel electrophoresis and hybridization with L. aethiopica kDNA. Control specimens from patients with skin diseases other than CL were all negative. Using PCR, Leishmania were demonstrated in the skin lesions of 7 cases in a group of 40 patients in whom the parasites could not be demonstrated by histopathology or culture in vitro although lesions were clinically suggestive of CL. These data indicate that PCR, carried out on DNA extracted from formalin-fixed and paraffin-embedded tissue specimens, is a valuable method for the diagnosis of CL, especially in chronic cases where the parasite load in the lesion is low.

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Newly-hatched chickens, isogenic for the B locus, were treated with high doses of cyclophosphamide, either alone or in combination with surgical bursectomy. The cyclophosphamide treatment initially caused virtual absence of bursal lymphoid cells and, later, complete destruction of the normal bursal architecture. It also caused an initial decrease in the lymphoid population of the thymus. However, thymic morphology was completely restored in chickens that were 15 days old or older. The most striking features in the morphology of the spleen and of the other peripheral lymphoid tissues of cyclophosphamide-treated birds was the absence of germinal centers and of the plasma cell line. No clear morphological differences could be detected between birds that were treated with cyclophosphamide alone and those subjected to cyclophosphamide treatment in combination with surgical bursectomy. The immunological capacities of normal, cyclophosphamide-treated and cyclophosphamide-treated bursectomized chickens were evaluated, starting with 1-month-old birds. The experimental groups of animals lacked or were profoundly deficient in agglutinating antibody to B. abortus antigen and to sheep erythrocytes after primary and secondary stimulation, while the normal controls responded well. Profound deficiency of IgM and IgG, as measured by the radial diffusion technique, was also obtained in the majority of the birds treated with cyclophosphamide alone or in combination with surgical bursectomy in the newly-hatched period. No clear differences could be detected, in the lack of capacity to respond to antigenic stimulation or to form immunoglobulins, between the birds that were treated with cyclophosphamide alone and those treated with cyclophosphamide together with bursectomy. Cellular immunological functions of normal and of cyclophosphamide-treated chickens were evaluated with regard to capacity to reject allografts and to elicit the graft-versus-host reaction. Cyclophosphamide treatment in the newly-hatched period did not affect the cellular immunological capacities of 1-month-old or older birds. Thus, by using this experimental protocol, cyclophosphamide can be used to accomplish ‘chemical bursectomy’, resulting in a permanent, severe deficiency of the humoral immunological capacities more frequently than can be obtained with other presently available bursectomy methods, while leaving the cellular immunological capacities intact.

Serum cytokine levels and mononuclear cell subpopulations in the spleen and peripheral blood of patients with visceral leishmaniasis before and after antimony therapy were analyzed. The percentages of activated monocytes/macrophages, T cells, and possibly B cells; of gamma/delta T cell receptor (TCR)-bearing T cells; of CD4- CD8- alpha/beta TCR-bearing T cells; and serum levels of tumor necrosis factor-alpha (TNF alpha), interferon-gamma (IFN-gamma), and interleukin-6 (IL-6) were high in patients with active visceral leishmaniasis. The proportion of both helper and suppressor CD4+ cells and of cells with NK and cytotoxic T phenotypes were depressed. Successful chemotherapy normalized these parameters with the exception of activated monocytes. Thus, the impaired cell-mediated immunity in human Leishmania donovani infection is primarily due to a decrease in the proportion and possibly the activity of helper CD4+ cells, while suppressor cells do not seem to play a relevant role. TNF alpha, IL-6, and IFN-gamma may prove to be useful markers for monitoring response to therapy.

The borough of Ocholo, on the western side of the Ethiopian Rift Valley, is an endemic focus for Leishmania aethiopica infection and has been surveyed thrice between 1987 and 1990. In 1989, 3022 inhabitants (> 95% of the population) were interviewed and examined. The overall prevalence of localized cutaneous leishmaniasis (LCL) was 3.6-4.0%, with a peak value of 8.5% in the 0-10 years old age group. In half of the patients the active disease was estimated to last for 9.6 +/- 6 months; in 10%, it exceeded 3 years. Scars of LCL were present in 34.3% of the residents. Leishmanin skin tests were positive in 54% of 120 school-children without signs of the disease. Therefore, in Ocholo a minimum of 71.6% of the population has been exposed to L. aethiopica infection. Two cases of the diffuse form of cutaneous leishmaniasis were observed. In this highland biotope, Phlebotomus pedifer was found to be the major, and possibly the only, vector for L. aethiopica.

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